Alzheimer's disease (AD) is an age-related neurodegenerative disease characterized by progressive cognitive decline and dementia. Our long-term goal is to understand the pathogenesis of AD and pave the way towards novel diagnostics and therapeutics. The objective here is to investigate the mechanistic link between dysregulation of mRNA splicing and neurodegeneration. Our central hypothesis is that AD-associated genetic variants influence mRNA splicing, which result in changes to pathways/networks and gene expression downstream contributing to cellular events leading to protein aggregation in AD. We will test our hypothesis by pursuing the following specific aims: In aim 1, we will perform a genome wide analysis using publically available multimodal expression data from post-mortem autopsy brain tissue (n=1,300) to discover aberrant RNA splicing events in the AD transcriptome. We will identify common and rare genetic variants regulating alternative splicing events in the brain (splicing QTLs). We will use novel long-read small molecule real time (SMRT) sequencing (IsoSeq) to generate full-length transcripts from 100 postmortem brain tissues to profile the full complexity of the AD transcriptome. We will validate these changes using proteomic, biochemical and cellular approaches. In aim 2, we will validate the RNA targets of disease-specific RNA binding proteins (RBPs) using enhanced crosslinking-immunoprecipitation (eCLIP) in control and AD brains. In aim 3, we will validate RNA targets of RBPs via lentivirally delivered shRNA knockdown and RBP binding sites via antisense oligonucleotide (ASO) inhibition in human pluripotent stem cell-derived neuronal and glial cell populations. This project will have a large overall impact by providing a comprehensive survey of RNA regulation in AD brain and adding functional and mechanistic interpretation of genetic variants associated with AD susceptibility. This contribution will be significant because it will provide a foundation for further mechanistic studies that will elucidate the drivers of disease and pave the way for novel therapeutic avenues, likely based on ASO- mediated approaches aimed at counteracting splicing changes by blocking proximal splicing enhancers or suppressors.